Clear cell carcinoma of the kidney (RCC), the most common form of kidney cancer, is associated with the inactivation of the von HippeI-Lindau (VHL) tumor suppressor. Mutations in the VHL gene can be found in approximately 70% of sporadic RCCs. One of the major functions of the VHL gene product, pVHL, is the targeting of the oxygen sensitive alpha-subunit of hypoxia-inducible factor (HIF) for ubiquitination and subsequent proteasomal degradation. Inactivation of VHL is felt to be an early event during RCC tumorigenesis and results in constitutive expression of two major HIF isoforms, HIF-1 and HIF-2. This results in increased transcription of genes that regulate glycolysis, angiogenesis and erythropoiesis. HIF-1 has furthermore been shown to up-regulate factors that promote growth arrest and apoptosis. The role of increased HIF expression in VHL associated tumorigenesis remains controversial. The hypothesis that the ratio of HIF-1 to HIF-2 levels is important for VHL associated renal tumor development will be investigated. Conditional gene targeting technology based on Cre-loxP mediated recombination as well as targeted transgenesis will be used to manipulate the expression levels of both pVHL and HIF in vivo and in vitro. This system enables the study of the functional relationship between VHL deficiency and HIF activation in regard to renal cell growth and viability in primary renal epithelial cells of different histogenetic origin. Specifically, the proposed investigations will examine the effects of increased or absent HIF-1 and increased HIF-2 expression in VHL deficient and wild type backgrounds. Studies will be performed in primary cell culture and in vivo with kidney specific conditional knock out mice. Gene expression studies will provide information regarding differential HIF-1 and HIF-2 target gene expression in different nephron segments with a special emphasis on genes involved in the regulation of cell survival. Taken all together, the proposed in vivo and in vitro studies will not only provide novel insights into the early events of renal oncogenesis and the histogenetic origin of RCC, but also examine fundamental aspects of HIF-1 and HIF-2 function in different nephron segments. Ultimately they have the potential to create a murine model of VHL associated renal tumors.